Neurotrophic factors are peptides that variously support the survival, proliferation, differentiation, size, and function of nerve cells (for review, see Loughlin and Fallon, Neurotrophic Factors, Academic Press, San Diego, Calif., 1993). While the numbers of identified trophic factors, or growth factors, are ever-increasing, most can be assigned to one or another established family based upon their structure or binding affinities. Growth factors from various families, including the epidermal growth factor (EGF) family, have been demonstrated to support dopaminergic neurons of the nigrostriatal system (an area that can be treated according to the methods of the present invention) (for review, see Hefti, J. Neurobiol. 25:1418-1435, 1994; Unsicker, Prog. Growth Factor Res. 5:73-87, 1994).
EGF, the founding member of the EGF family, was characterized more than 25 years ago (Savage and Cohen, J. Biol. Chem. 247:7609-7611, 1972; Savage et al., J. Biol. Chem. 247:7612-7621, 1972). Since then, additional members have been identified; they include vaccinia virus growth factor (VGF; Ventatesan et al., J. Virol. 44:637-646, 1982), myxomavirus growth factor (MGF; Upton et al., J. Virol. 61:1271-1275, 1987), Shope fibroma virus growth factor (SFGF; Chang et al., Mol. Cell. Biol. 7:535-540, 1987), amphiregulin (AR; Kimura et al., Nature 348:257-260, 1990), and heparin-binding EGF-like growth factor (HB-EGF; Higashiyama et al., Science 251:936-939, 1991). A common feature of these factors is an amino acid sequence containing six cysteines that form three disulfide cross links and support a conserved structure that underlies their common ability to bind the EGF receptor.
EGF is by far the most-studied member of the family and was the first localized to brain tissue: EGF-like immunoreactivity (IR) was found in areas of developing adult forebrain and midbrain including the globus pallidus, ventral pallidum, entopeduncular nucleus, substantia nigra, and the Islands of Calleja (Fallon et al., Science 224:1107-1109, 1984).
Another member of the EGF family, TGFα, has also been localized to brain tissue. It binds the EGF receptor (Todaro et al., Proc. Natl. Acad. Sci. USA 77:5258-5262, 1980), stimulates the receptor's tyrosine kinase activity, and elicits similar mitogenic responses in a wide variety of cell types (for review, see Derynck, Adv. Cancer Res. 58:27-52, 1992). TGFα might also bind to additional, unidentified receptors (which may help explain its differential actions in some cells). TGFα-IR has previously been shown to be heterogeneously distributed in neuronal perikarya throughout the adult rat CNS and in a subpopulation of forebrain astrocytes (Code et al., Brain Res. 421:401-405, 1987; Fallon et al., Growth Factors 2:241-250, 1990). TGFα mRNA has been detected in whole rodent brain (Lee et al., Mol. Cell. Biol. 5:3655-3646, 1985; Kudlow et al., J. Biol. Chem. 264:3880-3883, 1989) and in striatum and other brain regions by a nuclease protection assay (Weickert and Blum, Devel. Brain Res. 86:203-216, 1995) and by in situ nucleic acid hybridization (Seroogy et al., Neuroreport 6:105-108, 1994).
TGFα and EGF mRNAs reach their highest relative abundance (compared to total RNA) in the early postnatal period and decrease thereafter, suggesting a role in development (Lee et al., 1985, supra; Lazar and Blum, J. Neurosci. 12:1688-1697, 1992). In whole brain, the reduction is over 50% (Lazar and Blum, 1992, supra), whereas, in striatum, relative TGFα mRNA drops by two-thirds from peak levels (Weickert and Blum, 1995, supra). At all developmental stages examined, whole brain TGFα mRNA exceeds EGF mRNA levels by more than an order of magnitude (Lazar and Blum, 1992, supra).
The EGF receptor was localized by immunocytochemistry to astrocytes and subpopulations of cortical and cerebellar neurons in rat brain and to neurons in human autopsy brain specimens (Gomez-Pinilla et al., Brain Res. 438:385-390, 1988; Werner et al., J. Histochem. Cytochem. 36:81-86, 1988). EGF binding sites were revealed in rat cortical and subcortical areas, including the striatum, in an autoradiography study with radiolabeled EGF (Quirion et al., Synapse 2:212-218, 1988). In situ hybridization studies demonstrated EGF receptor mRNA in striatum and cells of the ventral mesencephalon (Seroogy et al., 1994, supra) and in proliferative regions in developing and adult rat brain (Seroogy et al., Brain Res. 670:157-164, 1995). As with relative EGF and TGFα mRNAs, EGF receptor mRNA is most abundant in striatum and ventral midbrain early in development, and gradually declines as the animal matures (Seroogy et al., 1994, supra).
Physiologically, TGFα acts on numerous cell types throughout the body, including many of neural origin (for review, see Derynck, 1992, supra). It supports the survival of cultured central neurons (Morrison et al., Science 238:72-75, 1987; Zhang et al., Cell. Regul. 1:511-521, 1990) and, unlike EGF, enhances survival and neurite outgrowth of dorsal root ganglion sensory neurons (Chalazonitis et al., J. Neurosci. 12:583-594, 1992). It also stimulates proliferation and differentiation of neuronal and glial progenitor cells from developing and adult brains (Anchan et al., Neuron 6:923-936, 1991).
The trophic effects of EGF-family peptides on mesencephalic dopaminergic neurons in culture have also been studied in recent years. EGF enhances the survival of E16 dopamine neurons in mixed midbrain cultures (Casper et al., J. Neurosci. Res. 30:372-381, 1991), but the degree to which it stimulates dopamine uptake is modest (Knusel et al., J. Neurosci. 10:558-570, 1990). TGFα also supports the survival of mesencephalic dopamine neurons in dissociated cell culture, but its effect is more selective than that of EGF (Ferrari et al., J. Neurosci. Res. 30:493-497, 1991; Alexi and Hefti, Neurosci. 55:903-918, 1993).
Another important characteristic of EGF-family growth factors is their ability to protect midbrain dopamine cells from neurotoxic assaults. EGF has been shown to protect dopamine neurons from glutamate or quisqualate excitotoxicity in dissociated cell culture (Casper and Blum, J. Neurochem. 65:1016-1026, 1995). It has also been demonstrated to protect cultured dopamine cells from the selective dopamine neurotoxin, 1-methyl-4-phenylpyridinium (MPP+; Park et al., Brain Res. 599:83-97, 1992) and to increase dopamine uptake in MPP+-treated cultures (Hadjiconstantinou et al., J. Neurochem. 57:479-482, 1991).
The results of studies with EGF in vivo were consistent those obtained in culture; EGF effected neuroprotection in both instances. For example, intracerebroventricular (ICV) infusions of EGF reduced amphetamine-induced rotations, increased the number of surviving tyrosine hydroxylase immunoreactive (TH-IR) cells in the SN, and increased striatal TH-IR fibers after transection of the nigrostriatal pathway in a rat model of PD (Pezzoli et al., Movement Disord. 6:281-287, 1993; Ventrella, J. Neurosurg. Sci. 37:1-8, 1993). ICV infusions of EGF into the brains of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesioned mice enhanced the content of dopamine and 3,4-dihydro-zyphenylacetic acid (DOPAC) and the activity of tyrosine hydroxylase in the striatum (Hadjiconstantinou et al., 1991, supra; Schneider et al., Brain Res. 674:260-264, 1995).
Despite its more potent activity in vitro, relative to EGF, the trophic effects of TGFα in vivo—particularly in animals, including humans, with neurological deficits—are undetermined. The present invention is based on newly discovered effects of TGFα infusion on cells in the normal and abnormal (lesioned) central nervous system, which are described herein.